The Applicant proposes to study microsomal electron transport in liver, kidney, and lung with specific emphasis on the physiological function(s) of NADPH-cytochrome c (P-450) reductase in its interaction with cytochrome P-450 and other electron carriers, such as cytochrome b5, and on the mechanism of electron transfer from the flavoprotein to various electron acceptors. Experimental approaches will attempt to: a. examine the electron transport systems in microsomal fractions from various organs with respect to the "specific" substrates involved; b. isolate and purify the NADPH-cytochrome c (P-450) reductases from these organs using the biospecific affinity chromatography procedure developed in the Principal Investigator's laboratory; c. isolate the cytochrome(s) P-450 from these organs using published procedures and other affinity chromatography methods presently being developed in the Principal Investigator's laboratory; d. determine the requirements for optimal interaction of the organ-specific reductase with its respective cytochrome P-450, including various phospholipids in the reaction mixtures or in liposomes constructed with a variety of lipid components; e. determine the nature of immunochemical similarities and differences among the reductases from various organs and species by electrophoretic mapping and amino acid analysis techniques; f. study the mechanism by which the NADPH-cytochrome c (P-450) reductase reduces various 1- and 2-electron acceptors, including its physiological acceptor, cytochrome P-450, and cytochrome b5; g. localize by immunohistochemical techniques, utilizing both light microscopy and electron microscopy, NADPH-cytochrome c (P-450) reductase in specific cells within various organs and in the endoplasmic reticulum of these cells.